Histological changes and neurotransmitter levels three months following perinatal asphyxia in the rat.

The involvement of excitatory amino acids (EAA) in the pathogenesis of hypoxic-ischemic states is well-documented. Information on the role of overexcitation by EAA in perinatalasphyxia (PA), however, is limited and data from adult models cannot be directly extrapolated to immature systems. Moreover, most adult models of ischemia are representing stroke rather than PA. We decided to study long term effects in a non-invasive rat model of PA resembling the clinical situation three months following the asphyctic insult. Morphometry on Nissl - stained sections was used to determine neuronal death in frontal cortex, striatum, hippocampus CA1, hypothalamus and cerebellum L1, and the amino acids glutamate, glutamine, aspartate, GABA, taurine, arginine as well as histamine, serotonin and 5-hydroxy-indoleacetic acid were determined in several brain regions and areas. Morphometry revealed that neuronal loss was present in the hippocampal area CA1 in all groups with PA and that morphological alterations were significantly higher in the cerebellar granular layer. The prominent light microscopical finding in all areas of asphyctic rats studied was decreased Nissl-staining, suggesting decreased cellular RNA levels. Glutamate, aspartate and glutamine were significantly elevated in the hypothalamus of asphyctic rats probably indicating overstimulation by EAA. Excitotoxicity in this area would be compatible with findings of emotional / behavioral deficits observed in a parallel study in our model of PA. Our observations point to and may help to explain behavioral and emotional deficits in Man with a history of perinatal asphyxia.

Evidence that taurine modulates osmoregulation by modification of osmolarity sensor protein ENVZ--expression.

Although the involvement of taurine in osmoregulation is well-documented and widely accepted, no detailed mechanism for this function has been reported so far. We used subtractive hybridization to study mRNA steady state levels of genes up- or downregulated by taurine. Rats were fed taurine 100 mg/kg body weight per day for a period of three days and hearts (total ventricular tissue) of experimental animals and controls were pooled and used for mRNA extraction. mRNAs from two groups were used for subtractive hybridization. Clones of the subtractive library were sequenced and the obtained sequences were identified by gen bank assignment. Two clones were found to contain sequences which could be assigned to the osmolarity sensor protein envZ, showing homologies of 61 and 65%. EnvZ is an inner membrane protein in bacteria, important for osmosensing and required for porine gene regulation. It undergoes autophosphorylation and subsequently phosphorylates OmpR, which in turn binds to the porine (outer membrane protein) promoters to regulate the expression of OmpF and OmpC, major outer membrane porines. This is the first report of an osmosensing mechanism in the mammalian system, which was described in bacteria only. Furthermore, we are assigning a tentative role for taurine in the osmoregulatory process by modifying the expression of the osmoregulatory sensor protein ENVZ.

Homocysteine increases cyclin-dependent kinase in aortic rat tissue.

BACKGROUND: Hyperhomocyst(e)inemia is strongly associated with occlusive arterial disease. A direct effect of homocysteine on the proliferation of smooth muscle cells was proposed recently. This observation led us to examine the effect of homocysteine on cyclin-dependent kinase, the starter of mitosis and reflecting proliferation. METHODS AND RESULTS: Seventy Him:OFA rats were divided into seven groups. For 12 weeks, 10 rats were fed homocysteine 25 mg/kg body weight per day, 10 were fed 50 mg/kg body wt per day, and 10 were fed 100 mg/kg body weight per day; 10 were given homocysteic acid 100 mg/kg body weight per day, 10 were administered cysteine 100 mg/kg body weight per day, and 10 were given ascorbic acid 270 mg/kg body weight per day. Ten remained untreated and served as controls. Aortic cyclin-dependent kinase was determined at the transcriptional (mRNA) and protein levels. Phosphokinase C and aortic homocyst(e)ine also were evaluated in aortic tissue. Aortic cyclin-dependent kinase protein was significantly (P = .0001) elevated in the three homocysteine-treated groups, and mRNA cyclin-dependent kinase levels were significantly elevated in the rats given the 50 and 100 mg/kg body weight per day protocol. Endothelial damage was shown at higher homocysteine doses as reflected by circulating ACE and von Willebrand factor changes. Proliferation of cells of the aortic wall by bromodeoxyuridine incorporation could be shown in the high-dose homocysteine group only. CONCLUSIONS: Our findings indicate that homocysteine specifically stimulates aortic cyclin-dependent kinase at the transcriptional level, with the possible consequence of proliferation of aortic cells as revealed by incorporation of bromodeoxyuridine in the aortic wall.

Decreased tumor incidence and increased survival by one year oral low dose arginine supplementation in the mouse.

The effects of arginine on tumor growth, antitumor mechanisms and a potential therapeutic role have been reviewed recently. In these studies, however controversial they were, high dose protocols for arginine treatment have been applied. Based upon own recent findings that low dose arginine stimulates the immune system and blocks lipid peroxidation, we performed preventive treatment with low dose (50 mg/kg body weight per day, orally administered) L-arginine in 150 mice for a period of one year. We compared survival and total number of tumors at the end of the feeding period to that found in 150 mice given taurine in the same dosage and in 150 mice without treatment. Survival of the arginine treated group was statistically significant as compared to that of the control group without treatment (p < 0.05): 116 mice were alive in the control group, 122 in the group administered taurine and 132 in the arginine treated group. The total number of tumors was significantly lower in the arginine treated group vs. the control group (p < 0.01). The total number of malign and benign tumors was significantly lower in the arginine treated group, whereas taurine significantly reduced the number of benign tumors only (p < 0.05). Arginine and taurine stimulate the immune system at the lymphocyte level. Arginine also acts at the macrophage level, inducing nitric oxide mediated cytotoxicity against tumor cells. Both compounds are known to block the formation of lipid peroxidation products. We therefore suggest that these two mechanisms are responsible for the decreased total number of tumors and the concomitant increase in survival.